Random access slide stainer with liquid waste segregation

ABSTRACT

An automated slide stainer with slides mounted in a horizontal position on a rotary carousel. Reagents and rinse liquids are automatically dispensed onto tissue sections or cells mounted on slides for the purpose of performing chemical or immunohistochemical stains. The rinse liquids are removed by an aspiration head connected to a source of vacuum. The aspiration head is a hollow chamber with a flattened bottom aspect. Eight holes in the bottom of the aspiration head provide for fluid communication between the exterior and interior of the aspiration head. To aspirate liquid off of a slide, the aspiration head is lowered so that the bottom aspect contacts the liquid. The liquid is sucked into the hollow interior of the aspiration head and collected into a liquid waste collection bottle. Several different liquid waste collection bottles are placed in parallel, between the source of vacuum and the aspiration head. A solenoid valve is positioned in line with each liquid waste collection bottle. Depending upon which solenoid valve is opened, the operator can direct the liquid waste to be collected into a specified liquid waste collection bottle.

BACKGROUND OF THE INVENTION

Tissue sections are commonly examined by microscopic examination, forboth research and clinical diagnostic purposes. Thin tissue sections orcellular preparations are commonly 1-10 microns thick, and are nearlytransparent if untreated. In order to visualize various histologicfeatures, a wide array of staining procedures have been developed overthe years that highlight various cellular or extracellular components ofthe tissues. Histochemical stains, also commonly termed "specialstains," employ chemical reactions to color various chemical moieties.Immunohistochemical stains employ antibodies as probes to color specificproteins, commonly via enzymatic deposition of a colored precipitate.Each of these histochemical and immunohistochemical stains requires theaddition and removal of reagents in a defined sequence for specific timeperiods. Therefore, a need arises for a slide stainer that can perform adiversity of stains simultaneously under computer control, as specifiedby the technologist.

Some of these histochemical and immunohistochemical stains employreagents that may be toxic, carcinogenic, or immiscible in water.Because of increasingly stringent local waste disposal requirements,many laboratories must now pay to dispose of these wastes throughspecial, hazardous, waste disposal companies. It is therefore desirableto minimize the volume of waste liquid that has to be treated ashazardous waste. With the advent of modern, sophisticated, slidestaining automation, it is therefore desirable to incorporate featuresinto an instrument to accomplish that task.

A common method for removing waste liquids from the surface of a slideis to flush or blow it off the surface of a slide into a common catchbasin. A representative example of such an approach is that described inU.S. Pat. No. 5,595,707. In that embodiment, reagent is removed from theslide by either blowing with a gas stream or flushing with a liquidreagent. Liquid falls off the slide into a catch basin. A similarapproach (using a common catch basin for all liquid waste) is taken inseveral other slide stainers, described in U.S. Pat. Nos. 5,425,918 and5,231,029, and that of Stark, E., et. al. 1988, An automated device forimmunocytochemistry, J. Immunol. Methods 107:89-92.

A similar design approach is evidenced by the slide stainer marketed byBioGenex Corporation, described in U.S. Pat. No. 5,439,649. It uses asimilar catch basin for collecting liquid waste. This design approachcauses the entire catch basin to become contaminated. The disadvantageof this approach is that the contaminated or toxic liquid is spread overa larger surface area than the slide itself, as it is caught in a basin.In order to ensure that the next waste liquid will not contain residualamounts of the toxic material, the catch basin must be flushed with alarger volume of wash solution. This results in an increased volume oftoxic liquid waste for special hazardous disposal.

An alternative design approach to handling waste liquids from a slidestainer is described in U.S. Pat. No. 4,543,236. Their invention shows ameans for aspirating liquid waste, under force of vacuum, to a commonwaste bottle. In that invention, liquid waste is aspirated through drainlines permanently connected to each slide-containing vessel. A dedicatedvalve for each slide-containing vessel opens to allow aspiration of theliquid contents. The system is "closed," in that the liquid supply andwaste lines do not become exposed to the atmosphere. An advantage ofthis approach is that liquid waste is not spread around a large catchbasin. However, the drawback of this design is that a dedicated valveand permanent tubing lines are required for each slide-containingvessel. As the number of slides increases, the apparatus becomesexpensive and complex to assemble and repair. This limitation wasapparent in their particular embodiment, as the staining apparatus onlyaccommodated five slides.

A conceptually similar approach is described in U.S. Pat. No. 4,358,470,except that in this invention, waste liquids are channeled into theiroriginal containers, to be used repetitively. Their invention did notrequire a large number of distinct procedures to be applied to differentmicroscope slides. Rather, all of the biological specimens, mounted onelectron microscopic grids, were held in a common chamber and treated inan identical manner. With only a single incubation chamber, permanentlyclosed plumbing lines for liquid supply and waste was a reasonable,cost-efficient, design. It would not be applicable to situations wherenumerous slides are to be stained using different chemical stainingprocedures.

A third method of automated slide staining for immunohistochemistry wasdescribed by Brigati, U.S. Pat. No. 4,731,335. In that invention, liquidwas applied to and removed from capillary gaps that were formed by twoslides closely apposed together. To remove the liquid, the edges of theslides were abutted against absorbent towels, causing the liquid to beadsorbed. Therefore, waste liquid was in a solid, adsorbed form.

A fourth method for rinsing slides has been to simply dip the slidescontaining a reagent into a vat of liquid, such as water or buffer. Thereagent dilutes out in the excess volume of liquid wash solution,preparing the slide for treatment with the next reagent that isscheduled to be applied. An example of that approach is the slidestainer described in U.S. Pat. No. 4,092,952. A similar (dipping ofslides into a vat) approach, specially tailored forimmunohistochemistry, is described in a publication by Muir andAlexander, 1987, Easier immunoperoxidase staining with labour savingincubator box. J Cain Pathol 40:348-50.

A previous invention by one of the present inventors, U.S. Pat. Nos.4,847,208 and 5,073,504, disclosed a means for aspirating liquids fromthe surface of slides. A pipette is manually lowered until it is incontact with the wetted slide surface. Liquid was aspirated into asingle waste bottle by vacuum force.

SUMMARY OF THE INVENTION

This invention relates to an improved slide staining device, for theapplication and removal of reagents to biologic tissue sections mountedon microscope slides. The improvement relates to a method of segregatingliquid wastes after application to biological specimens mounted onmicroscope slides. With this invention, it is possible to remove liquidsfrom the microscope slide surface and collect some waste liquids indifferent containers than other liquids. Certain waste liquids, such asorganic solvents, liquids immiscible with water, or biohazardouschemicals, are not to be flushed down the drain in many cities. Rather,local water resource regulations require that these compounds besegregated from regular aqueous waste, and disposed of through specialmethods. Moreover, the invention incorporates a novel aspiration headthat efficiently removes liquid from the entire surface of themicroscope slide. This invention provides a means to collect toxic wasteliquids in small volumes for economical disposal.

In accordance with the present invention, a slide stainer comprises aslide support adapted to support at least one microscope slide in ahorizontal position to retain liquid on its surface. An aspiration head,in fluid continuity with the source of vacuum, is caused by an actuatorto contact the liquid on the slide surface. A liquid director directsliquid waste to collect in a selected one of plural waste collectioncontainers.

Preferably, the aspiration head comprises a hollow manifold havingplural apertures through a planar surface which is essentially parallelto the microscopic slide during liquid waste aspiration. The planarsurface contacts the liquid on the slide but does not directly touch thebiological specimen.

Preferably, plural slides are mounted in a horizontal position on arotary carousel and a liquid aspiration station is provided in a fixedlocation on the periphery of a carousel, the carousel being moved toselect the slide from which liquid is aspirated.

More specifically, the preferred embodiment comprises a rotary carouselof microscope slides carrying biologic samples, such as tissue sectionsor cell smears. The slides are indexed to a liquid aspiration stationwhich has an aspiration head for removing the liquid waste from thesurface of the microscope slide. Since the liquid is spread out in aplanar fashion, upon a flat microscope slide, the aspiration head isdesigned to have a similarly shaped flat bottom surface. Eight holes arepresent in the bottom surface of the aspiration head that allow forcommunication between the hollow aspiration head and the exterior. Thehollow interior is in fluid continuity with a source of vacuum. Severalliquid waste bottles are positioned in a parallel configuration, betweenthe vacuum source and the aspiration head. Each liquid waste bottleinlet is normally closed off with a solenoid valve. When liquid is to beaspirated, a selected bottle's solenoid valve opens. The aspiration headis electromechanically lowered so that it's bottom surface contacts theliquid on the microscope slide. In this manner, suction force istransmitted directly to the holes on the aspiration head, causing theliquid to be collected in the selected liquid waste bottle.

BRIEF DESCRIPTION OF THE DRAWINGS

The foregoing and other objects, features and advantages of theinvention will be apparent from the following more particulardescription of preferred embodiments of the invention, as illustrated inthe accompanying drawings in which like reference characters refer tothe same parts throughout the different views. The drawings are notnecessarily to scale, emphasis instead being placed upon illustratingthe principles of the invention.

FIG. 1 is a perspective view of a first embodiment of a slide stainer.

FIG. 2 is a top view of a slide frame for providing five sealed cavitiesabove five different slides holding tissue samples.

FIG. 3 is a top view of a slide frame base.

FIG. 4 is a bottom view of a slide frame housing.

FIG. 5 is a top view of the slide frame housing with five microscopeslides in their appropriate positions, showing the area to which heat isapplied.

FIG. 6 is a cross-sectional view of a slide frame resting on the sliderotor.

FIG. 7 is a schematic diagram of the heater and sensor wiring diagram,on the slide frame, and the interconnection with the temperaturecontroller.

FIG. 8 is a side cross-sectional view of a cartridge pump dispensingmechanism in the liquid dispensing and removal station.

FIG. 9 is a side cross-sectional view of a bulk liquid dispensingstation housed in the liquid dispensing and removal station.

FIGS. 10A and 10B are side cross sectional views of a vacuum hose andtransport mechanism for removing liquid reagent and wash fluids fromslides contained on the slide rotor.

FIG. 11A is a side cross-sectional view of the aspiration head, showingits relationship to the glass slide in the slide frame.

FIG. 11B is a bottom end face view of the aspiration head.

FIG. 12 is a perspective view of a second embodiment of a slide stainer.

FIG. 13 is a perspective view of the liquid handling zone of the secondembodiment of the slide stainer.

FIGS. 14A and 14B are side cross-sectional views of the liquidaspiration station of the second embodiment, with the aspiration head inthe lowered (FIG. 14A) and raised (FIG. 14B) positions.

FIG. 15 is a schematic representation of the waste liquid pathways ofthe second embodiment.

FIG. 16 is a schematic representation of the bulk liquid dispensepathways of the second embodiment.

FIG. 17 is a schematic representation of the individual heaters on theslide rotor and the temperature control boards mounted on the sliderotor.

FIGS. 18A-D are a schematic diagram of the electronic circuitry of thetemperature control board.

DETAILED DESCRIPTION OF THE INVENTION

FIG. 1 shows a first embodiment 1 of the invention in perspective view.Generally, the first embodiment 1 comprises a substantially circularassembly base 2, a slide rotor 3 rotatable on the assembly base 2, areagent rotor 4 also rotatable on the assembly base, and a liquiddispensing and removal station 5.

The slide rotor 3 is driven to rotate by a servo motor (not shown) andcarries ten slide frames 6 that are radially asserted into anddetachable from it. A top view of single slide frame 6 is shown in FIG.2. Here, positions for five slides, each with a tissue sample, are shownin positions 7a-7e. The slide frame 6 comprises a slide frame base 8shown in FIG. 3. The slide frame base 8 includes a heated area 9 whichunderlies each of the slide positions 7a-7e and incorporates resistiveheating elements, not shown. The heating elements are integrally formedin the slide frame base 8. Electricity for powering the heating elementsis provided into the slide frame 6 from the assembly base 2 via firstand second contacts 10. Further, third and fourth contacts 11 enabletemperature sensing of the heated areas via thermocouples alsointegrally formed in the slide frame base 8. In practice, a sum of threeconnectors are required, since contacts 10 and 11 share the same groundconnection. Therefore, one of the connectors 11 are left unused.

Adapted to overlay the slide frame base is a slide frame housing 12.FIG. 4 is a top view of the slide frame housing 12 showing essentially arigid plastic or metal frame 13 with five oval holes 14a-14ecorresponding to each of the slide positions 7a-7e. A silicon rubbergasket 15 is also provided under the frame 13. Returning to FIG. 2, theslide frame housing 12, including the gasket 15 and frame 13, is boltedonto the slide frame base 8 by two Allen bolts 16 to provide individualsealed cavities approximately 0.2-0.4 inches deep over each tissuesample slide placed at each of the slide positions 7a-7e. As a result, atotal of 3 ml of reagents and/or rinses can be placed in contact withthe tissue samples of each one of the slides but a maximum quantity of 2ml is preferable. Since the silicon gasket 15 is compressed by the frame13 against the microscope slides (not shown), the cavities over each ofthe frame positions are mutually sealed from each other.

FIG. 5 is a top view of a slide frame base 8 with five microscope slides17 in the positions denoted by 7a-7e in FIG. 3. The area of each slide17 forming cavities, that are delimited by the silicone rubber gasket 15and holes 14a-14e is indicated by an approximately rectangular line 18,marking the chamber wall. The area denoted by the hatched bars indicatesthe area of the slide frame base 8 that includes heating elements 9. Theentire heated area (hatched bars) is raised to the same temperature,bringing the group of five slides to the same desired temperature. Theportion of each slide 17 that is not above the heated area does notgenerally bear a biologic tissue specimen. Rather, it is used forlabeling purposes.

FIG. 6 is a cross-sectional view of an assembled slide frame base 8 andhousing 12, collectively referred to previously as the slide frame 6.The microscope slide 17 is shown held in position, between the slideframe base 8 and housing 12. The slide frame 6 is resting on the sliderotor 3. In this view, the electrical connection between the slide frame6 and an edge connector 19 is demonstrated. Four edge connectors perslide frame 6 are provided (contacts 10 and 11 in FIGS. 2 and 3). Theelectrical connection is fed from the edge connector 19 through theslide rotor via an insulated feed-through 20, to a terminal underneaththe slide rotor 3. A wire then connects the terminal to a source ofpower or control circuitry (not shown).

FIG. 7 is a schematic diagram, showing two out of the ten heater 91 andsensor 92 circuits that can be placed on the instrument slide rotor. Theheater is represented schematically as a resistive element, andcorresponds to the heated area (hatched bars) of FIG. 5. Contacts 10 and11 share a common ground connection, leaving one of the four connectorsunused. Each of the circuits feeds into a temperature controller,represented schematically 21. Each slide frame sends three wires to thetemperature controller 21--a heater power conductor 22, a sensorconductor 23, and a ground connection 24. The temperature controller 21is mounted in a stationary position on the assembly base 2. Since theheaters and sensors are in frequent motion, they connect to thestationary temperature controller 21 via a service loop (not shown). Theservice loop contains the wires from each of the edge connectors 19.Sufficient extra length is provided in the wires so that as the sliderotor rotates, the service loop travels around the slide rotor axis. Theslide rotor 3 does not turn more than one full revolution in eitherdirection. The wires in the service loop are preferably bundled togetherwith a wire tie, so that individual wires do not become entangled orcaught underneath the slide rotor 3. Since there are three wires percircuit (wires 22-24), and there are ten slide frames 6 on the sliderotor 3, the service loop contains a minimum of thirty wires.

Referring to FIG. 1, positioned above the slide rotor 3 is the reagentrotor 4. This reagent rotor is similarly adapted to rotate on theassembly base 2 and is driven by another servo motor (not shown) undercomputer control (not shown). The reagent rotor 4 and the slide rotor 3rotate independently of each other. The reagent rotor 4 is adapted tocarry up to ten cartridge frames 25. Each of these cartridge frames 25are detachable from the reagent rotor 4 and can be selectively attachedat any one of ten possible points of connection. Each cartridge frame 25is capable of carrying five of the cartridge pumps 46.

Generally, the dispensing station 5 comprises a soft hammer 26 forengaging a portion of the cartridge pumps 46. The cartridge pumps 46 areconstructed so as to dispense liquid when a portion of the cartridgepump 46, called the metering chamber 42 of the cartridge pump 46 iscompressed. It is possible to dispense from any of a plurality ofcartridge pumps by rotating the reagent rotor so as to align a desiredcartridge pump 46 with the hammer 26. This provides the capability ofdispensing precisely measured amounts of reagent to any slide positionedunderneath the cartridge pump 46 adjacent to actuator 26. The mechanismfor dispensing from the cartridge pumps 46 is shown in greater detail inFIG. 8. The hammer 26 is driven by a solenoid or linear stepping motor43 that is mounted on a front wall 44, attached to the assembly base 2.In FIG. 8, the hammer is shown compressing the metering chamber 42portion of the cartridge pump. It is important to be able to adjust thespeed of compression by the hammer 26 upon the metering chamber 42.Otherwise, too rapid a compression will cause an excessively forcefulejection of reagent from metering chamber 42, potentially damaging thetissue section underneath. Therefore, a linear stepping motor ispreferred instead of a solenoid. As another alternative, thereciprocating hammer of the dispensing actuator could take the form of acam, driven by a rotary motor, that engages the metering chamber 42 sothat the rotation of the cam will compress the metering chamber.

The cartridge pump 46 is comprised of a liquid reservoir 45 and themetering chamber 42. The liquid reservoir 45 shown in this firstembodiment 1 is a syringe barrel. The metering chamber 42 is comprisedof a compressible elastomeric housing with a one-way inlet valve (notshown) and a one-way outlet valve (not shown), both valves aligned in adownwards direction of fluid flow. When the hammer 26 compresses themetering chamber 42, the liquid reagent contained within is ejected.When the compressive force is removed, the negative pressure created bythe expansion of the elastomeric housing, trying to resume its native,non-compressed shape, causes liquid to flow inwards from the liquidreservoir 45. In this manner, repetitive compression of the meteringchamber 42 causes repetitive dispensing of small aliquots of reagent.Alternative cartridge pumps are presented in U.S. patent applicationSer. No. 08/887,178 filed Jul. 2, 1997 and U.S. patent application Ser.No 09/020,983 filed Feb. 10, 1998, which are incorporated herein byreference.

The dispensing station 5 further includes a means to dispense liquidsfrom a large bottle (FIG. 9). Bulk liquid bottles 27 that can supplyliquid into any one of the microscope slides 17 on any one of the slideframes 6 via rinse tubes 28. Each bulk liquid bottle 27 is connected toits own rinse tube 28. The bulk liquid bottles 27 are pressurized by apump (not shown). The outflow tube (not shown) from each bulk liquidbottle 27 passes through a valve 47 that regulates the flow of liquidfrom that bottle. By opening the valve for a defined period of time,under computer control (not shown), with a defined pressure within thebottle 27, a known quantity of liquid can be dispensed onto the slide17. The liquids placed within the bottles 27 are those that are usedrepeatedly among many different procedures, such as water, saline, andalcohol.

As shown in FIG. 9, the bulk liquid bottles 27 are screwed into a femalethreaded cap 48 secured to the underside of the horizontal top wall 49of the station frame. Compressed air from a compressor (not shown) isprovided to each bulk liquid bottle 27 through a pressure regulator 50.Tubing from the pressure regulator 51 transmits the compressed air tothe inlet of the bulk liquid bottle 27. The pressure above the liquidenables the liquid to forced up through the dip tube 52 through therinse hose 53 when a pinch valve 47 is opened. Depending on the lengthof time that the pinch valve is opened, a pre-determined amount ofliquid can be dispensed through the rinse tube 28.

The liquid dispensing and removal assembly 5 further includes a liquidremoval vacuum station, positioned adjacent to the rinse tubes 28 (notvisible in FIG. 1). In order to remove liquid from the surface of aslide 17, the reagent rotor positions the slide at the liquid removalvacuum station, shown in a side cross-sectional representation in FIGS.10A and 10B. An external source of vacuum (not shown) is channeledthrough a trap flask 29, ultimately leading to a vacuum hose 30 thatterminates in an aspiration head 31. The tubing connections are notshown in FIGS. 10A and 10B. The vacuum hose 30 and aspiration head 31are supported by a hose transport mechanism 54 that allows theaspiration head 31 to be extended down into a cavity of a slide frame 6to remove liquid covering the tissue sample on the slide 17. As theaspiration head contacts the liquid, the liquid is sucked upwards intothe tubing and collected into the trap flask 29.

The vacuum hose transport mechanism 54 comprises a motor 32. Areciprocating link 33 is attached to a crank arm 34 so that the rotationof the motor 32 causes the reciprocating link 33 to traverse in avertical direction. A bottom portion of the reciprocating link 33 isconnected to a lever 55 that is pivotally attached to the station frame.The other end of this lever is connected to a vacuum hose clamp 35 thatis connected via pivot arms 36 to a plate 37 rigidly attached to thestation frame. The net effect of these connections is that when themotor 32 is rotated, the slide arm 33 descends in a vertical direction.Thus, the lever 55 is pivoted clockwise around its fulcrum causing thehose clamp 35 to pivot up and away on the two pivot arms 36 from theslide as shown in FIG. 10B. The motor is automatically turned off as thelink 33 reaches its two extreme ends of movement by the contact of theelectrical terminals 39 of the link to the contact plates 38 connectedto the station frame.

The aspiration head 31 is shown in greater detail in FIGS. 11A and 11B.FIG. 11A shows the aspiration head in a lowered position, incross-section, within the cavity formed by the slide frame 6. Theaspiration head 31 comprises a hollow interior manifold 40 through whichthe vacuum force is transmitted across the entire lower surface of theaspiration head 31. Eight holes 41 are drilled on the lower face of theaspiration head 31, through which the suction force is transmitted.Since the microscope slide 17 is planar, liquid on the slide surfacespreads out in two dimensions. Therefore, in order to thoroughly removeliquid from all portions of the microscope slide 17, multiple aspirationsites are needed. We accomplish this with an aspiration head with aplanar lower surface with multiple holes. The planar surface of theaspiration head 31 comes into close parallel apposition to themicroscope slide 17. The aspiration head only contacts the liquid, notthe microscope slide itself, lest it damage the glass slide 17 or thebiologic specimen that it carries (not shown). Without such a design andonly a single aspiration site, such as from a pipette, liquid distantfrom the aspirator would not be removed. Rather, it would cling to thedistant surfaces of the glass slide 17, because of the surface tensionon the glass. This would result in a residual volume of liquid thatwould otherwise be left on the surface of the slide 17. Having a closeparallel apposition of the aspiration head is also helpful from theperspective of decreasing surface tension during liquid aspiration. Theclose parallel apposition of the bottom surface of the aspiration headwith the microscope slide 17 creates a type of capillary gap. This gaphelps to overcome surface tension, ensuring complete liquid removal.

A computer (controlled 86 in FIG. 5) controls the instrument functions.That is, an operator programs the computer with the information such asthe location of reagents on the reagent rotor and the location of slideson the slide rotor. The operator then programs the particularhistochemical protocol to be performed on the tissue samples. Variablesin these protocols can include the particular reagent used on the tissuesample, the time that the tissue sample is allowed to react with thereagent, whether the tissue sample is then heated, the rinse that isthen used to wash the reagent away, followed by the subsequent removalof the rinse and reagent to allow subsequent exposure to a possiblydifferent reagent. The instrument enables complete random access, i.e.,any reagent to any slide in any sequence.

A second, preferred, embodiment of the invention is shown in FIG. 12.Like the previous embodiment, it also comprises two independentcarousels that rotate on an assembly base 56. Bulk liquid bottles 57 aremounted on a bridge 58 that extends across the width of the entiremachine, above the reagent rotor. A separate group of trap bottles 59,for collecting waste liquid, are mounted on the side of the bridge 58 ina compartmentalized shelf. The tubing connections and valves for thebulk liquid bottles 57 and the trap bottles 59 are hidden from view byan upper panel 60. The front and sides of this embodiment are surroundedby a plexiglass case 61, that can be manually slid sideways in order toinsert cartridge pumps 62 or slides (not shown). Slides are individuallyinserted and removed via a centrally located slide access door 63. Theslides (not shown) are hidden from view by a circular platen 64 that islocated above the slides and reagent rotor (not shown). Functionssimilar to the dispensing assembly (5 of FIG. 1) in the previousembodiment are accomplished in a somewhat similar liquid handlingassembly (not shown) that is positioned in a liquid handling zone 65.

FIG. 13 shows the individual mechanisms contained within the liquidhandling zone 65, including a hammer 66 for dispensing from cartridgepumps (not shown), an aspiration head 67 for removing liquid from thesurface of slides, a bulk liquid dispensing port 68, and an air-mix head69 for spreading and mixing liquids on the surface of a slide. Theelectromechanical mechanism for dispensing from cartridge pumps, bycompressing a hammer 66 upon a metering chamber of a cartridge pump (notshown in FIG. 13), is similar to the previous embodiment (FIG. 8).Reagent dispensed from the cartridge pump (not shown) flows onto theslide by passing through a roughly rectangular hole in the platen 64.

The aspiration head 67 also functions in a similar manner to that of theprevious embodiment. In order to simplify the linkage mechanism forlowering and raising the head 67, the head moves solely in a verticaldirection. This is shown in further detail in FIGS. 14A and 14B. FIG.14A shows a side cross-sectional view of the aspiration head in a downposition, within a cavity formed by the microscope slide 75 (bottomsurface) and a slide chamber clip 76 (lateral walls). As in the firstembodiment, a gasket (not shown) seals the surface where the slidechamber clip 76 contacts the microscope slide 75. A linear stepper motor73 moves the aspiration head up and down, under computer control(demonstrated schematically in FIG. 15). As in the first embodiment 1,the aspiration head 67 comprises a hollow manifold 74 connected to asource of vacuum. Eight holes communicate between the bottom of theaspiration head 67 and the exterior, through which liquid is aspirated.When vacuum is supplied to the aspiration head 67, and the head 67 islowered adjacent to the slide, the liquid reagent on top of the slide isaspirated off and collected in a trap bottle 59 (shown schematically inFIG. 15). When the aspiration head 67 is not in use, it is raised to theup position (FIG. 14B), allowing free rotation of the slide rotor 77.

FIGS. 14A and 14B also show the physical location of a heating element78, represented as a resistive element inside a rectangular box withcross-hatched lines. Each slide rests directly on the heating element78, so that heat is directly communicated to the microscope slide. Athermistor is incorporated into each heating element (not shown in FIGS.14A and 14B). Each of forty-nine microscope slides 75 has its ownheating element 78, so that the temperature of each slide 75 can beindependently regulated. Power for the heating element 78 is supplieddirectly from a temperature control board 79 that is affixed to theunderside of the slide rotor 77. Seven identical temperature controlboards 79 are so mounted underneath the slide rotor 77, evenly spacedaround the periphery. Each temperature control board supplies power forseven heating elements 78. The means by which this is accomplished isexplained later, in reference to FIGS. 17 and 18A-D.

An important aspect of this embodiment is the provision for thesegregation of waste liquids that are removed from the surface of theslide. A schematic diagram explaining how this is accomplished is shownin FIG. 15. Three different waste bottles 59 are mounted on theinstrument. Connections 70 are also provided on the instrument for alarge external trap bottle 71, typically of a ten or twenty litercapacity for aqueous waste. Four solenoid valves, labelled 80A-80Dcontrol to which bottle aspirated liquid will be directed. These valvesare under computer control, schematically represented by the boxlabelled "controller" 86. Valve 81 is a three way valve. It can allow adirect connection between the vacuum pump 82 and the overflow trap 83,or between the pump and the ambient environment. A connection to theambient environment is required if the aspiration system needs to bebypassed when the air-mix head 69 is in use. If valves 80A and 81 areappropriately opened, the pump 82 turned on, and the aspirator head 67lowered so as to aspirate liquid, the liquid will be directed upwardsinto the tubing, as represented by the arrow "fluid flow." Liquid willthen follow the only path available, and be collected into the externaltrap bottle 71. Valves 80B-80D function similarly for their respectivetrap bottles 59. A small overflow trap bottle 83 is also inserted intothe line with its own fluid sensor 93. This provision is included so asto detect if any of the trap bottles 59, or external trap bottle 71 areoverflowing with waste liquid. In that case, liquid would enter theoverflow trap bottle and be detected by the fluid sensor. Thatinformation would be communicated to the controller 86, which would shutthe system down and alert the instrument operator on the computerscreen.

Referring to FIG. 13, the liquid handling zone also includes an air-mixhead 69. A schematic representation of the air flow into the air-mixhead 69 is shown in FIG. 15. The pump generates a high velocity airstream that is channeled into the air-mix head 69. Air intake to thepump is via the three way solenoid valve 81 (FIG. 15). The solenoidvalve 81 (FIG. 15) switches so as to channel air directly from theatmosphere to the pump (FIG. 15), bypassing the aspiration system andtrap bottles 59 and 71. The high velocity air flow is focused onto theslide. The air-mix head 69 travels back and forth along the length ofthe slide, pushed and pulled by a belt and pulley that is attached to amotor (not shown). The net effect of this system is to direct a curtainof air back and forth along the length of the slide, causing liquid tobe mixed and spread along the surface of the microscope slide.

The liquid handling zone 65 (FIG. 12) includes a bulk liquid dispensingport 68 (FIG. 13). The function of the rinse tubes 28 of the firstembodiment 1 (shown in FIG. 1) are all incorporated into a single bulkliquid dispensing port 68 in this preferred embodiment. Therefore,slides are positioned under the bulk liquid dispensing port 68regardless of the bulk liquid bottle that the liquid is actually derivedfrom. A schematic representation of the fluid pathways and controlvalves is shown in FIG. 16. The bulk liquid bottles 57 are eachconnected to a source of pressure, that is generated by a pump 85. Thepressure is communicated to the bulk liquid bottles 57 via a pressuremanifold 94. Solenoid valves 72a-72f are placed between the bulk liquiddispensing port 68 and each bulk liquid bottle 57. Liquid flows out thebulk liquid dispensing port 68 only when one or more of the valves72a-72f are open. A pressure switch 84 also communicates with thepressure manifold 94. It is capable of sensing the amount or pressurecontained within the manifold 94. When it falls below a specified level,it communicates with the controller 86 causing activation of the pump85. As the pump generates an increased amount of air pressure within thepressure manifold, the pressure switch resets, causing the pump to stoppumping. In this manner, a relatively constant pressure head ismaintained within the pressure manifold 94.

A dispense sensor 95 is positioned underneath the bulk liquid dispensingport 68 to provide verification that liquid was dispensed when one ofthe solenoid valves 72a-72f were transiently opened. The dispense sensor95 comprises an optical sensor and an LED light source. When liquid isdispensed from the bulk liquid dispensing port 68, the liquid interruptsthe light beam. The change in resistance across the sensor as a resultof the decrement in light intensity is communicated to the controller86.

This second, preferred embodiment of the invention includes thecapability to independently heat the forty-nine slides to differenttemperatures. A novel aspect of this embodiment is the method forindependently regulating the amount of power that each of the forty-nineheaters receives. Moreover, each heater also incorporates a temperaturesensor. Each of these sensors must communicate with the computer 86 inorder to allow for appropriate temperature feedback and regulation. Inthe first embodiment 1, groups of up to five slides were under a single,common temperature control mechanism. Each heating group had wires thatdirectly connected with the temperature controller (FIG. 7). With threewires per group (power for heat, sensor feedback, and a shared ground)and ten groups of slides, at least thirty wires were contained in theservice loop. If a similar system were used for forty-nine differentheaters, as in this preferred embodiment, 147 wires would be required inthe service loop. Such a bulky service loop would be problematic.Therefore, an alternative method is developed in this preferredembodiment.

FIG. 17 shows the relationship between each of the heating elements 78mounted on the slide rotor 77, depicting the heating element 78 as aresistive element. A single sensor 87 is adjacent to each heater. Thecombination of a single heating element 78 and sensor 87 are sopositioned so as to provide a location 88 for a single slide to beheated. The physical layout of this location 88 is demonstrated in FIGS.14A and 14B. Two wire leads from each heating element 78, and two wireleads from each sensor 87 are connected directly to a temperaturecontrol board mounted on the slide rotor 77. Each temperature controlboard is capable of connecting to up to eight different heater andsensor pairs. Since this embodiment incorporates forty-nine slidepositions, seven boards 79 are mounted to the underside of the sliderotor, each connecting to seven heater-sensor pairs. One heater-sensorposition per temperature controller board 79 is not used. Also shown inFIG. 17 is the serial connection 89 of each of the seven temperaturecontrol boards, in a daisy-chain configuration, by six wires. The firsttemperature control board is connected via a service loop 90 to thecomputer 86 (FIG. 16) which serves as the user interface and systemcontroller. The service loop contains only six wires.

FIG. 18 is an electronic schematic diagram of the temperature controlboard 79. The design of the temperature control board 79 was driven bythe need to minimize the number of wires in the flexible cable (serviceloop 90) between the heaters and the computer. To minimize the length ofwires, seven temperature controller boards 79 are used, each mounted onthe slide rotor. Thus, each heater is positioned close to its associatedelectronics and the size of each board 79 is kept small because eachruns only seven heating elements 78. Each temperature controller board79 includes the function of an encoder and decoder of temperature data.That data relates to the actual and desired temperature of each ofheating elements 78. The data flows back and forth between the computer86 and the temperature control board 79. If an individual heatingelement 79 requires more or less heat, the computer communicates thatinformation to the temperature control board 79. The temperature controlboard 79, in turn, directly regulates the amount of power flowing toeach heater. By placing some of the logic circuitry on the slide rotor,in the form of the temperature control boards 79, the number of wires inthe service loop 90, and their length, are minimized.

In this embodiment, the temperature control board 79 system was designedas a shift register. The machine's controlling microprocessor placesbits of data one at a time on a transmission line, and toggles a clockline for each bit. This causes data to be sent through two shiftregister chips U1 and U2 on each control board, each taking eight bits.There are thus 16×7 or 112 bits to be sent out. Referring to FIGS.18A-D, the data comes in on connector J9.1, and the clock line is J9.2.The shift registers used in this design are "double buffered," whichmeans that the output data will not change until there is a transitionon a second clock (R clock), which comes in on pin J9.3. The two clocksare sent to all seven boards in parallel, while the data passes throughthe shift register chips (U1 and U2) on each board and is sent on fromthe second shift register's "serial out" pin SDOUT to the input pin ofthe next board in daisy chain fashion. It will be seen that a matchingconnector, J10, is wired in parallel with J9 with the exception ofpin 1. J10 is the "output" connector, which attaches via a short cableto J9 of the next board in line, for a total of seven boards. The otherthree pins of J9 are used for power to run the electronics (J9.4),electronic ground (J9.5), and a common return line (J9.6) fortemperature measurement function from the sensors.

Of the sixteen data bits sent to each board, eight from register U2control the on/off status of up to eight heating elements 78 directly.This can be accomplished with a single chip because shift register U2has internal power transistors driving its output pins, each capable ofcontrolling high power loads directly. Four of the remaining eight bitsare unused. The other four bits are used to select one thermistor 87 outof the machine's total complement of forty-nine. For reasons of economyand to reduce the amount of wiring, the instrument has only oneanalog-to-digital converter for reading the forty-nine temperaturetransducers (thermistors 87), and only one wire carrying data to thatconverter. This channel must therefore be shared between all of thetransducers (thermistors 87), with the output of one of them beingselected at a time. Component U4 is an analog multiplexer which performsthis function. Of the four digital bits which are received serially, oneis used to enable U4, and the other three are used to select one of thecomponent's eight channels (of which only seven are used). If pin fouris driven low, U4 for that board 79 becomes active and places thevoltage from one of the seven channels of that board on the sharedoutput line at J9.6. Conversely, if pin four is pulled high, U4's outputremains in a high impedance state and the output line is not driven.This allows data from a selected board 79 to be read, with the remainingboards 79 having no effect on the signal. Multiplexer U4 can only beenabled on one board 79 at a time; if more than one were turned on at atime, the signals would conflict and no useful data would betransmitted.

Temperature sensing is accomplished by a voltage divider technique. Athermistor 87 and a fixed resistor (5.6 kilohms, R1-R8, contained inRS1) are placed in series across the 5 volt electronic power supply.When the thermistor is heated, its resistance drops and the voltage atthe junction point with the 5.6 kilohm resistor will drop.

There are several advantages to the design used in this embodiment.Namely, the temperature control boards 79 are small and inexpensive.Moreover, the heater boards are all identical. No "address" needs to beset for each board 79. Lastly, the service loop 90 is small in size.

An alternative potential design is that each temperature control board79 could be set up with a permanent "address" formed by adding jumperwires or traces cut on the board. The processor would send out a packetof data which would contain an address segment and a data segment, andthe data would be loaded to the board whose address matched the addresssent out. This approach takes less time to send data to a particularboard, but the address comparison takes extra hardware. It also demandsextra service loop wires to carry the data (if sent in parallel) or anextra shift register chip if the address is sent serially. Yet anotherpotential design is that each temperature control board 79 could haveits own microprocessor. They could all be connected via a serial datalink to the main computer 86. This approach uses even fewer connectingwires than the present embodiment, but the cost of hardware is high. Italso still implies an addressing scheme, meaning that the boards wouldnot be identical. Also, code for the microprocessors would be required.

Equivalents

While this invention has been particularly shown and described withreferences to preferred embodiments thereof, it will be understood bythose skilled in the art that various changes in form and details may bemade therein without departing from the spirit and scope of theinvention as defined by the appended claims. Those skilled in the artwill recognize or be able to ascertain using no more than routineexperimentation, many equivalents to the specific embodiments of theinvention described specifically herein. Such equivalents are intendedto be encompassed in the scope of the claims.

What is claimed is:
 1. A slide stainer comprising:a slide supportadapted to support a plurality of microscope slides in a horizontalposition to retain liquid on the surface of each; an aspiration head influid continuity with a source of vacuum; an actuator which causes theaspiration head to contact the liquid on the slide surface but notcontact the slide surface; a controller and electromechanical means formoving said aspiration head relative to said plurality of slides so asto position said aspiration head immediately above a selected slide fromwhich liquid is to be aspirated; and a liquid director which directsliquid waste to collect in a liquid waste collection container.
 2. Aslide stainer as claimed in claim 1, wherein the surface of theaspiration head that contacts the liquid on the slide is essentiallyplanar.
 3. A slide stainer as claimed in claim 2, wherein the aspirationhead has more than one aperture through which to aspirate liquid.
 4. Aslide stainer as claimed in claim 2, wherein the planar surface isconnected to a mechanical linkage capable of positioning said planarsurface in contact with the liquid on the slide but not in contact withthe biological specimen.
 5. A slide stainer as claimed in claim 2,wherein the planar surface is essentially parallel to the microscopeslide during liquid waste aspiration.
 6. A slide stainer as claimed inclaim 1, wherein the aspiration head comprises a hollow manifold havinga plurality of apertures through which to aspirate liquid.
 7. A slidestainer as claimed in claim 1, wherein a plurality of slides are mountedon a rotary carousel and a liquid aspiration station comprising theaspiration head is provided in a fixed location on the periphery of saidrotary carousel.
 8. A slide stainer as claimed in claim 1 wherein theliquid director directs liquid waste to collect in one of plural liquidwaste collection containers.